Fig. 5
GSK2795039 improves neuronal survival in co-culture with activated microglia. a Primary neurons were co-cultured with BV2 microglia exposed to FeSO4, DMSO, LPS, and/or GSK2795039. Immunocytochemistry was completed for neurons (NeuN) at 24 h after co-culture. All groups were counter-stained with DAPI. Qualitatively, a reduction in the expression of NeuN was noted on those groups co-cultured with microglia treated with FeSO4, LPS, or LPS&FeSO4. The addition of GSK2795039 significantly improved neuronal survival. b Improved survival among all cell types present in the neuronal culture (DAPI-positive) was observed as increased cell counts in groups treated with GSK2795039. c Quantitative analysis of NeuN-positive cell counts revealed a significant reduction in neuronal cell count after co-culture with microglia treated with FeSO4 & LPS & DMSO. GSK2795039 prevented neuronal loss after co-culture and significantly improved neuronal cell counts. d PC12 cells were differentiated to a neuronal like phenotype for one week prior to microglia co-culture. Immunocytochemistry revealed increased staining for cleaved caspase-3 among PC12 co-cultured with BV2 microglia treated with FeSO4, LPS, or LPS&FeSO4. The addition of GSK2795039 to the microglia treatment groups reduced the presence of cells positive for cleaved caspase-3. e Quantitative analysis of cleaved caspase-3 positive PC12 cells showed significant increase when co-cultured with microglia treated with FeSO4 & LPS & DMSO. However, this increased apoptosis among PC12 cells was reduced by the addition of GSK2795039. All immunocytochemistry images are representative of one trial. All groups were compared using one-way ANOVA with Tukey’s post-hoc test. In all graphs, symbols representing significance were assigned according to comparisons: DMSO group (*) and FeSO4 & LPS & DMSO ($). *p < 0.05, **p < 0.01, ***p < 0.001, ****p < 0.0001, $$p< 0.01, $$$p< 0.001, $$$$p< 0.0001. All graphs represent n = 4. Bars represent mean ± SEM. Size bars represent 50 μm